Project 1 will characterize and define synaptic and cellular phenotypes of high-risk mutations associated with schizophrenia (SCZ) (NRXN1 exonic deletions, 22q11.2 deletions and 16p11.2 duplications), in human induced neuronal (iN) cells derived from iPS cells. We will identify the most robust phenotypic changes in mutated iN cells (engineered and naturally-occurring), and the electrophysiological, genomic and morphometric assays that detect those differences most cost-effectively. The most promising models and assays will be selected for future high-throughput screening, based on robustness (prioritizing those models and assays that reveal phenotypes that are observed in more than one mutation) and cross-validation across laboratories, within and across species, and across source tissues in mice (iN cells, primary cultured neurons and mPFC brain slices -- Project 2). The project includes 9 specific aims: (1) To generate iN cells with an engineered form of each high-risk mutation for functional evaluation of mutant and non-mutant cells. (2) To generate iN cells from iPS cells from patients carrying high-risk mutations and controls. (3) To characterize the synaptic phenotype(s) of these mutations in human iN cells using electrophysiology and functional imaging. (4) To identify novel morphological synaptic and cellular phenotypes of these mutations in iN cells using morphometric analysis of high-definition images. (5) To identify gene regulatory networks associated with high-risk mutations. (6) To test cross-lab reproducibility of all procedures and findings. (7) To integrate functional, molecular and morphological data from mouse and human, mutant and control iN cells. In this last Aim, which is ongoing throughout the study, taking into account all results from Aims 3-6 in human iN cells as well as from Project 2 (mouse models), the most robust and reproducible pathophysiological models will be identified; the extent to which synaptic and cellular phenotypes are overlapping across mutations vs. distinct will be evaluated; and recommendations will be made for the selection of one or more model systems and assays for future high- throughput screening of novel therapeutics. This work will proceed through the analysis of each of the three mutations of interest, studying iN cells first in the engineered form of the mutation compared with non-mutant cells from the same control line, and then in the naturally-occurring mutations observed in SCZ patients vs. control individuals. This work will develop one or more pathophysiological models of synaptic and cellular dysfunction in iN cells carrying specific mutations, for future use in high-throughput screening of novel therapeutics.